The objectives of this research are to determine the chemical mechanisms of the concerted interactions of several intermediary metabolites with single allosteric enzymes, and the relationship of the structure of these enzymes to their function. The enzyme chosen for these studies its the phosphoenolpyruvate carboxylase of Escherichia coli. This is an allosteric enzyme that plays a major role in the control of the flow of intermediary metabolites through the citric acid cycle. Some of the compounds that stimulate its catalytic activity in vitro are, acetyl coenzyme A, some nucleotide di- and triphosphates, long chain fatty acids and their coenzyme A derivatives, fructose-1,6-bisphosphate, and some organic solvents. The most effective inhibitors of this enzyme's activity are four-carbon dicarboxylic acids that contain a nucleophilic group in the L-alpha-configuration, e.g., aspartate, malate. The mechanisms of the interactions of these effectors on the enzyme, singly and in combinations, are studied primarily by kinetic methods, but augumented by other physical chemical methods where feasible. Partially active and inactive enzyme preparations obtained from mutants of E. coli are used to identify which amino acid changes are responsible for the altered functions. The amino acid changes are determined by conventional methods on protein fragments obtained by specific enzymatic digestion or specific chemical cleavage. Attempts are made to obtain altered enzymes that are no longer capable of forming higher order aggregates in vitro so that relative importance of subunit-subunit interactions in the regulation and catalytic functions of this enzyme can be determined.